Taxol stabilizes microtubules and is one of the most effective drugs for the treatment of metastatic breast and ovarian cancer, yet the mechanism by which taxol induces growth arrest and cytotoxicity is not well understood. In studying taxol's mechanism of action after the drug binds to microtubules, we have shown that taxol induces, in a dose-and time- dependent fashion, accumulation of the cyclin inhibitor p21 in both p53 wild type and p53-null cells. In cells expressing wild type p53, this protein is also induced by taxol treatment, and this induction is mediated primarily by increased stability of the protein. Coincident with these effects, taxol treatment results in activation of c-raf-1 kinase and MAP kinase. Pharmacologic depletion of c-raf-1 with benzoquinone ansamycins abrogates both the ability of taxol to induce both wild type p53 and the cyclin inhibitor p21, as well as taxol-induced cytotoxicity, implicating c-raf-1 as a mediator of taxol action. Raf-l has been shown to form a molecular complex with the anti-apoptotic protein Bcl-2. Following taxol treatment, this complex dissociates, coincident with the phosphorylation of Bcl-2. Bcl-2 phosphorylation has been associated with its inactivation. Since pharmacologic depletion of c-raf-1 also blocks Bcl-2 phosphorylation following taxol, our data implicate c-raf-1 as the kinase responsible for inactivating Bcl-2 following taxol treatment. Our findings suggest a novel mechanism of taxol-induced cytotoxicity, involving a distinct subcellular pool of c- raf-l, and involving Bcl-2 and the cyclin inhibitor p2l.